Evidence of In Vivo Existence of Borrelia Biofilm in Borrelial Lymphocytomas.

Lyme borreliosis, caused by the spirochete Borrelia burgdorferi sensu lato, has grown into a major public health problem. We recently identified a novel morphological form of B. burgdorferi, called biofilm, a structure that is well known to be highly resistant to antibiotics. However, there is no evidence of the existence of Borrelia biofilm in vivo; therefore, the main goal of this study was to determine the presence of Borrelia biofilm in infected human skin tissues. Archived skin biopsy tissues from borrelial lymphocytomas (BL) were reexamined for the presence of B. burgdorferi sensu lato using Borrelia-specific immunohistochemical staining (IHC), fluorescent in situ hybridization, combined fluorescent in situ hybridization (FISH)-IHC, polymerase chain reaction (PCR), and fluorescent and atomic force microscopy methods. Our morphological and histological analyses showed that significant amounts of Borrelia-positive spirochetes and aggregates exist in the BL tissues. Analyzing structures positive for Borrelia showed that aggregates, but not spirochetes, expressed biofilm markers such as protective layers of different mucopolysaccharides, especially alginate. Atomic force microscopy revealed additional hallmark biofilm features of the Borrelia/alginate-positive aggregates such as inside channels and surface protrusions. In summary, this is the first study that demonstrates the presence of Borrelia biofilm in human infected skin tissues.

Effectiveness of Stevia Rebaudiana Whole Leaf Extract Against the Various Morphological Forms of Borrelia Burgdorferi in Vitro.

Lyme disease is a tick-borne multisystemic disease caused by Borrelia burgdorferi. Administering antibiotics is the primary treatment for this disease; however, relapse often occurs when antibiotic treatment is discontinued. The reason for relapse remains unknown, but recent studies suggested the possibilities of the presence of antibiotic resistant Borrelia persister cells and biofilms. In this study, we evaluated the effectiveness of whole leaf Stevia extract against B. burgdorferi spirochetes, persisters, and biofilm forms in vitro. The susceptibility of the different forms was evaluated by various quantitative techniques in addition to different microscopy methods. The effectiveness of Stevia was compared to doxycycline, cefoperazone, daptomycin, and their combinations. Our results demonstrated that Stevia had significant effect in eliminating B. burgdorferi spirochetes and persisters. Subculture experiments with Stevia and antibiotics treated cells were established for 7 and 14 days yielding, no and 10% viable cells, respectively compared to the above-mentioned antibiotics and antibiotic combination. When Stevia and the three antibiotics were tested against attached biofilms, Stevia significantly reduced B. burgdorferi forms. Results from this study suggest that a natural product such as Stevia leaf extract could be considered as an effective agent against B. burgdorferi.

Short-term electrical stimulation enhances the effectiveness of Botulinum toxin in the treatment of lower limb spasticity in hemiparetic patients.

The study tested the spasmolytic effect of Botulinum toxin A in two groups of hemiparetic patients with lower limb spasticity: in the first group (n = 5) 2000 U Dysport were injected into the soleus, tibialis posterior and both heads of gastrocnemius muscles alone; the second (n = 5) received additional repetitive alternating electrical stimulation of M. tibialis anterior and plantar flexors for 30 min six times per day during the 3 days following the injection. Muscle tone, rated by the Ashworth spasticity score, and gait analysis including recording of vertical ground reaction forces, were assessed before and 4 weeks after injection. The combined treatment proved to be more effective with respect to the clinically assessed reduction of muscle tone, gait velocity, stride length, stance- and swing-symmetry (P < 0.05). The result is discussed with reference to animal experiments demonstrating enhanced toxin uptake and accelerated onset of its paralytic effect by electrical stimulation.

Gait function in spastic hemiparetic patients walking barefoot, with firm shoes, and with ankle-foot orthosis.

The purpose of the study was to investigate the effect of firm shoes and a particular type of a frequently prescribed ankle-foot orthosis in Germany, the Valens calliper, on functional gait parameters in hemiparetic patients with marked extensor spasticity and corresponding heel-strike problems. Cycle times, vertical ground reaction forces, trajectories of the centre of pressure (gait line) under the soles and the mode of initial contact were assessed in 19 hemiparetic patients walking barefoot, with firm shoes and the orthosis. With the shoes, significant improvements (p < 0.006) were observed for: walking velocity, stride length, initial double stance duration and gait line of the non-affected foot. The pattern of the vertical force diagram improved qualitatively in six patients. With the orthosis the following variables showed an even more marked improvement (p < 0.006): walking velocity, cadence, stride length, and gait line of both the affected and non-affected foot. Fourteen patients became rear-foot strikers and the vertical force pattern further improved in six patients. It is concluded that the ankle-foot orthosis (more than the shoes) provide a useful support in the early rehabilitation of hemiparetic patients with marked spasticity.

Ankle muscle activity before and after botulinum toxin therapy for lower limb extensor spasticity in chronic hemiparetic patients.

BACKGROUND AND PURPOSE: Recent studies have been made of the novel treatment of lower limb spasticity after stroke with botulinum toxin A, and the results were based mostly on the clinical assessment made before and after treatment. This study investigated the effects of toxin on ankle muscle activity during gait in patients with severe extensor spasticity. The questions posed were whether the toxin particularly diminishes the so-called premature muscle activity as a major cause of equinovarus deformity and whether different types of altered motor control allow a prediction of the outcome of the treatment. METHODS: In 12 chronic hemiparetic outpatients with pronounced lower limb spasticity, we injected 400 U botulinum toxin A into the soleus and tibialis posterior muscles and both heads of the gastrocnemius muscles. Ankle spasticity and complex gait analysis including kinematic electromyography (EMG) of the soleus and tibialis muscles were assessed before treatment and 4 weeks after the injection. RESULTS: Nine patients profited with a reduction of spasticity, improved gait ability, and a more normal temporal pattern of muscle activity with a prominent reduction of the premature activity of the plantar flexors. Eight patients exhibited a qualitative pattern (type I) corresponding to an increased stretch-reflex excitability. Three patients did not profit; their muscle tone, gait ability, and muscle activation remained stable or even deteriorated. CONCLUSIONS: This study further supports the beneficial effects of botulinum toxin in the treatment of lower limb extensor spasticity. A correlation was observed between the clinical reduction of muscle tone, functional gait parameters, and a more normal EMG pattern with a predominant reduction of the premature activity of the plantar flexors. The qualitative type of EMG pattern corresponding to an increased stretch-reflex excitability (type I) was a positive predictor for the outcome.

Synergism of penicillin and gentamicin against Listeria monocytogenes in ex vivo hemodialysis culture.

A new technique of ex vivo hemodialysis culture was used to study the effects against Listeria monocytogenes of penicillin G and gentamicin administered intra-muscular, singly and in combination, in the goat. Bactericidal synergism of the combination was detected despite rapidly diminishing concentrations of the drugs in the blood plasma and dialysate.